Accumulation of brain protein aggregates is a hallmark of Alzheimer?s disease and many other neurodegenerative disorders. Brain protein aggregation was originally assumed to be a cell-autonomous phenomenon in Alzheimer?s disease, but increasing evidence indicates that non?cell-autonomous processes also play major roles by promoting the spread of toxic aggregation-prone proteins. Although the principal components of the brain protein aggregates that accumulate in Alzheimer?s disease victims have been detected in extracellular vesicles (EVs), the mechanisms underlying the spread of protein aggregates in Alzheimer?s disease are poorly understood. Recent work in our lab suggests that the GBA gene plays an important role in this process. GBA encodes the lysosomal enzyme glucocerebrosidase, which catalyzes the conversion of the sphingolipid glucosylceramide to glucose and ceramide. Mutations in GBA cause several neurodegenerative diseases characterized by the accumulation of protein aggregates in the brain. To study the mechanisms by which mutations in GBA cause disease, we created a Drosophila model of glucocerebrosidase deficiency by deleting the Drosophila GBA ortholog, GBA1b. GBA1b mutants accumulate ubiquitinated protein aggregates, show age-related neurodegeneration, have changes in the turnover and abundance of EV proteins and exhibit a six-fold elevation in EV abundance. Furthermore, ectopic expression of GBA1b in peripheral tissues such as muscle or gut rescued protein aggregation in the brain. These findings lead us to hypothesize that mutations in GBA1b result in lipidomic alterations promoting the overproduction of extracellular vesicles that spread protein aggregates from peripheral tissues to the brain and possibly between cells in the brain. To test this hypothesis and explore the underlying mechanisms, we propose three aims. First, we will test whether the non?cell-autonomous rescue of brain protein aggregation in GBA1b mutants is mediated by extracellular vesicles, and whether this pathway influences the protein aggregates seen in other common neurodegenerative diseases, including Alzheimer?s disease. Second, we will test whether GBA1b mutants promote the spread of endogenous protein aggregates, as well as those seen in Alzheimer?s and prion disease, by blocking the production of EVs in GBA1b mutants and by transplanting EVs from GBA1b mutants to WT flies. Third, we will perform proteomic, lipidomic and cell biological experiments to explore how mutations in GBA1b alter extracellular vesicle formation and composition. Given the increasing evidence of peripheral influences on the spread of brain protein aggregates in Alzheimer?s disease and other neurodegenerative disorders, we anticipate that our findings will advance our understanding of this phenomenon and also create novel opportunities for therapeutic intervention in these diseases.